Reaction vessel for receiving minimal quantities of fluid samples

ABSTRACT

In accordance with the invention, a method for executing reactions at temperatures exceeding 50° C. is disclosed using minimal quantities of fluid samples. The method uses a reaction vessel which minimizes the containment space for the fluid sample. The reaction vessel includes a sample-receiving tube and an insert. The insert engages the sample-receiving tube, forming a seal and prevents evaporation of the fluid sample during heating. The sample-receiving tube of the method is tapered to be narrow towards the bottom so that the insert can be easily inserted into the sample-receiving tube from the top and provide a tight seal in the lower extremity without increasing pressure in the reaction chamber. Equalization between the two sides of the sealing insert can occur as the insert is being introduced. Minimal displacement of the insert in the axial direction of the sample-receiving tube produces a sealing connection minimizing undesirable pressure build up.

BACKGROUND OF THE INVENTION

The invention relates to a reaction vessel for receiving minimalquantities of fluid samples.

In particular, molecularbiological work often calls for reactions attemperatures exceeding 50° C. (thru 100° C.) as in the high-temperatureinactivation of proteins, denaturing nuclein acids, restrictionaldigestion with Taq I and the like. These reactions are usually carriedout in standard reaction vessels placed in preheated water baths or inholes drilled heated metal blocks. The volumes of the reaction solutionsare normally in the range 10 and 50 μl, the volumes of the reactionvessels between 1000 and 2500 μl. The reaction vessels thus contain alarge excess volume, in which water evaporates from the reactionsolutions and is deposited on the inside of the cover. This increasesthe concentrations in the reaction solution, sometimes to the extentthat the sample completely dries out. This is greatly obstructive to thetests and treatments involved and can only be avoided by specialmeasures such as, for instance, repeated centrifuging of the reactionvessels or coating the reaction solution with oil.

OBJECT OF THE INVENTION

The object of the invention is to create a reaction vessel of theaforementioned type in which the gas volume in a reaction vessel filledwith a reaction solution is reduced to such an extent that evaporationof the reaction solution into larger spaces located above is no longerpossible, thus effectively preventing drying out of the sample in heattreatment.

One aspect of the invention is to reduce the volume for the sample inthe reaction vessel with an insert to such an extent that the space forevaporation above the sample is configured as small as possible toaccordingly limitate evaporation of the fluid from the sample.

A further aspect of the invention is to configure a seal active on allsides by arranging said insert as tight as possible above the samplelocated in the sample-receiving tube so that the remaining volume abovethe sealed off area is isolated gas-tight from the actual reactionchamber in which the sample is located. In this way any resulting vaporis restricted to the resulting, comparitively small reaction chamber.

A substantial problem is posed by configuring the seal between theinsert and the sample-receiving tube so that when the insert is pushedinto the sample-receiving tube little or no pressure is built up in thereaction chamber. This is important, for one thing, because otherwisepushing the insert into the sample-receiving tube is only possible byconsiderable exertion, and, for another thing, buildup of pressure inthe reaction chamber could be detrimental to the sample or to the teststo be carried out.

To permit isolating a relatively small reaction chamber in the area ofthe lower extremity of the sample-receiving tube without any tangiblepressure build-up in the reaction chamber when pushing the insert intothe sample-receiving tube a seal is formed only after the insert is atits terminal position.

Since the cross-section of the sample-receiving tube is larger above thesealing area in the lower extremity than at this extremity, the insertcan be pushed into the sample-receiving tube from the top by its sealinglower extremity or by the sealing element provided at its lowerextremity without causing pressure to build up, because a gap permittingpressure equalization remains between the periphery of the sealing areaor sealing element and the internal wall of the sample-receiving tube.It is not until the sealing area or the sealing element comes intocontact with the area of the reduced cross-section that the sealingconnection is momentarily produced, i.e. with minimal displacement ofthe insert in the axial direction. Thus, there is practically nobuild-up of pressure within the reaction chamber. Whilst the insert maybe formed very voluminous which is particularly useful in the embodimenthaving no seal at the lower extremity of the insert, an embodimentemploying a relatively thin rod is preferred because here very littlematerial is required for the insert, it merely serving as a carrier andpressure transfer element for the sealing element provided at its lowerextremity, said sealing element having sealing contact with the interiorwall of the sample-receiving tube.

By releasably attaching the insert to the lid, the separate reactionvessel can be used either with or without insert, as required.

The insert can also be replaced so that, for example, inserts ofdiffering length and with differing sealing elements at the lowerextremity can be used, with which reaction chambers of differing sizecan be separated from the remaining volume.

Particularly when sealing the reaction chamber in the area of the lowerextremity of the insert, venting of the lid is useful to prevent anybuild-up of pressure in the remaining volume.

BRIEF DESCRIPTION OF THE DRAWINGS

The invention will now be described by way of example with reference tothe following Figures, wherein:

FIG. 1 is vertical, longitudinal section through the center of areaction vessel according to a first embodiment of the invention,

FIG. 2 is a corresponding section through the lower extremity of areaction vessel according to the invention featuring a slightlydifferent configuration of the seal,

FIG. 3 is a section, similar to FIG. 2, of an embodiment operating witha graduated annular seal,

FIG. 4 is a section, similar to FIG. 1, of an embodiment of the reactionvessel featuring a rod-type insert, and

FIG. 5 is a section through the lower extremity of a reaction vesselaccording to FIG. 4 showing a different embodiment of the sealingelements.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT

According to FIG. 1 sample-receiving tube 11 having a circularcross-section features above a rounded tip 24 a conical section 11""extending upwards, followed by a relatively short cylindrical section11"', topped by a short extension section 11" and completed by a topcylindrical section 11' which is relatively long and has the largestdiameter relative to the other sections.

At the top of the exterior wall of the sample-receiving tube 11a malethread 26 designed to mate with the female thread 27 of the screw-on lid12 is provided. On its inside the lid 12 has a circular-cylindricalrecess 28 which is open towards the bottom, located concentrically tothe centerline 23 and in which a hollow-cylindrical insert 14 isinserted from underneath as a sliding fit, or better, as a seized fit.Basically the insert 14 could just as well be secured in the recess 28also by using a suitable fastener such as an adhesive, for example.However, the insert 14 is preferably provided for withdrawing when thelid 12 is unscrewed, thus facilitated replacement.

The circumferential flange 29 forming the recess 28 features a radiallyoutwards directed slight spacing 30 from the interior wall 15 of thesample receiving tube 11 so that between the top end of the wall 31 anda bottom annular wall 32 of lid 12 when screwed fully in place aclearance 33 remains which borders on a side vent opening 25 in thethreaded circumferential edge 34 of the lid 12. In this way pressurecompensation is possible between the interior space 13 of thesample-receiving tube 11 and atmosphere.

A hollow cylindrical insert 14 has a slightly smaller outer diameterthan the circular-cylindrical section 11"' and changes at its lowerextremity into a extremity section 20 which is slightly conical in taperdownwards, at the lower extremity of which the insert 14 is closed offby a bottom wall 17.

The conical extremity section 20 has a seal contact 35 with the interiorwall 15 of the sample-receiving tube 11 within the section 11"" when lid12 is screwed in place as shown in FIG. 1 , said interior wall 15 alsobeing tapered downwards in this section. In this way the extremitysection 20 together with the floor wall 17 at the lower extremity of thesample-receiving tube 11 separates a reaction chamber 19 in which asample 18 is filled, which is to be exposed to heat treatment by placingthe sample-receiving tube 11 in a heating apparatus.

It is assumed that in the reaction chamber 19 a quantity of approx. 100μl of sample is introduced. This quantity of sample almost fills thereaction chamber 19 completely so that when heat is applied theresulting vapor is forced to remain practically completely within thesample 18 thus preventing it from drying out.

Should only minor quantities of the sample 18--for instance only 50μl--require treatment, the insert 14 could be replaced by a somewhatlonger and more tapered insert 14 as indicated in FIG. 1 by the dashedline. In this way a much smaller reaction chamber 19' could be separatedfrom the overall internal volume of the sample-receiving tube 11.

Due to the insert 14 being replaceable one-and-the-same lid 12 could befurnished with inserts 14 of differing length for separating reactionchambers of differing volume.

On the outer circumference of the screw cap 12 a fluted surface 21 isprovided to facilitate unscrewing and screwing into place. A furtherfluted surface 36 is provided on the outer circumference of the topsection 11' of the sample-receiving tube 11 to present added resistancefor the other hand too, when unscrewing.

Functioning of the reaction vessel as described above is as follows:

With the lid 12 unscrewed and the insert 14 removed the reaction vesselfirst receives the sample 18 in the desired quantity in thesample-receiving tube 11. Then, depending on the quantity of samplefilled, an insert 14 of suitable length is selected and inserted fromunderneath into the screw cap 12. The insert 14 is then introduced intothe sample-receiving tube 11 so that the lid 12 can then be screwed intoplace on the male thread 26, during which the lid 12 must have adequatefreedom of movement in axial direction so that the conical extremitysection 20 of the insert 14 contacts the internal wall of the conicalsection (or part) 11"" of the sample-receiving tube 11 and by turningthe lid 12 further in the closing direction sufficient axial force canbe produced in the direction of the arrow via the underside 37 of thelid 12 and the top edge 38 of the insert 14 to create the seal 35.

As soon as the seal 35 is produced a certain residual clearance shouldremain at 33.

In a top recess 39 of the lid 12 a marking label 40 is inserted.

In the embodiment as shown in FIG. 2 contact is made not by the completebottom conical extremity section 20 of insert 14 with the conicalinternal wall of section 11"" but merely by a lip 20' provided in thecircumference of the bottom wall 17.

Since there is a distinct space between both the extremity section 20and the circumferential lip 20' and the internal wall 15 when the insert14 is introduced from above until it comes into contact with saidinternal wall 15, the reaction chamber 19 can be vented immediatelyprior to the seal 35 is produced thus preventing a substantial pressurebuild-up in the reaction chamber 19 when producing the seal 35.

In the embodiment as depicted in FIG. 3 an annular step 22 is providedat the bottom extremity of the circular-cylindrical sector 11"'projecting inwards which acts together with the conical extremitysection 20 of the insert 14 in a stuffing effect, by means of which too,the reaction chamber 19 can be separated from the remaining volumelocated above gas-tight.

In all embodiment examples the reference numbers refer to correspondingcomponents.

The embodiment example as shown in FIG. 4 differs from that of FIG. 1 inthat instead of the hollow cylindrical insert 14 a relatively thin,rod-shaped insert 14' of solid material is provided, having at itsbottom extremity a sealing plate 17' which produces a seal contact 35with the interior wall of the conical section 11"" when the lid 12 isscrewed into place. The rod-shaped insert 14' can also be separated fromthe lid 12 when withdrawn from the latter and, for example, be replacedby a longer or shorter insert 14'. Whilst in the embodiment exampleaccording to FIG. 1 venting is provided by a lateral borehole 25 in thelid 12, the embodiment example according to FIG. 4 a groove 25' ismachined in the female thread 26, said groove providing venting of theinternal space 13 via the clearance 33 and the circumferential gap 30.

Otherwise, the function is the same as in the embodiment as shown inFIG. 1. In FIG. 5 an annular step 22 can be provided--also inconjunction with a rod-shaped insert 14'--at the bottom extremity of thecircular cylindrical sector 11"', said step acting together with theplug 20" secured to the bottom extremity of said rod-shaped insert 14'.

We claim:
 1. A method for reacting a given volume of a fluid sample at atemperature at which the fluid sample generates evaporation, the methodcomprising the steps of providing an elongated sample tube having aninterior wall, a first, closed end, a second, open end and a sample tubevolume, placing a given volume of fluid sample into the closed end ofthe tube, inserting an insert having an end portion capable of sealingsaid sample tube volume wherein proper alignment and such that aremainder of the insert projects from the tube and moving the endportion towards the closed end of the tube, venting a space inside thetube between the closed end and the end portion to an exterior of thetube while moving the end portion during the moving step until the endportion is in a position relative to the tube at which the closed end ofthe tube and a surface of the end portion proximate the closed end ofthe tube define a space, establishing a fluid tight, circumferentialseal between the end portion of the insert and the interior wall of thetube when the end portion is at said position, thereafter holding theend portion and the tube in their relative positions in which theydefine said space, and reacting the fluid sample at said temperatureduring the holding step, whereby an escape of evaporation generatedduring the reacting step from said space is prevented.
 2. A methodaccording to claim 1 wherein the step of venting comprises the step ofmaintaining an air passage between the insert, including the end portionthereof, and the interior wall of the tube prior to the step ofestablishing the fluid-tight, circumferential seal.
 3. A methodaccording to claim 1 including the step of closing the open end of thetube when performing the step of establishing the fluid-tight,circumferential seal.
 4. A method according to claim 3 including thestep of passing air at least during the closing step through a passagein a lid or between the lid and the interior wall of the tube.
 5. Amethod according to claim 4 wherein the step of passing comprisesmaintaining said passage open after the closing step has been completed.6. A method according to claim 5 wherein the step of establishing thecircumferential seal includes the step of forming opposing sealingsurfaces on the interior wall and the end portion which are activated bymoving the sealing surfaces against each other in an axial direction ofthe tube, and including the step of exerting an axially-oriented forceagainst the circumferential seal to increase the sealing effectestablished thereby.
 7. A method according to claim 6 wherein the stepof exerting comprises the step of exerting the axially-oriented force onthe end portion with the lid for the tube connected with the endportion.